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BACKGROUND: Tissue engineering has become a major field of research in biotechnology and biomedicine. As a consequence, cell-based therapeutic approaches are entering the hospitals, especially for skeletal regeneration. Traumatic injuries of cartilage are treated with autologous cell suspensions or in vitro generated cartilage tissues, but there is actually no therapy available for degenerative cartilage defects. However, Osteoarthritis (OA) is a major public health problem in the world affecting 240 million people globally.
OBJECTIVE: To develop suitable in vitro tissues, the properties of chondrogenic spheroids should be optimized via coculture with cells naturally occurring as joint neighbours.
METHODS: Human chondrocytes were isolated from condyles and propagated in monolayer culture. Scaffold-free spheroids were generated and co-cultured with joint-specific partner cells (osteoblast-like osteosarcoma cells, fibroblasts). Morphology and differentiation was analyzed using histochemistry (Alcian blue, Safranin O) and immunohistochemistry for cartilage markers (collagen type II, Sox9, proteoglycan), proliferation-associated protein (Ki67) and markers of connective tissue (collagen type I and actin).
RESULTS: The provision of a more natural microenvironment in vitro via co-culture of chondrocyte-based aggregates with osteoblast-like Saos-2 cells enhanced the differentiation potential of chondrogenic spheroids towards hyaline cartilage.
CONCLUSIONS: The study showed the positive influence of Saos-2 cells on the differentiation potential of human chondrocytes in co-culture.
Lens epithelium-derived growth factor splice variant of 75 kDa (LEDGF/p75) is an autoantigen over-expressed in solid tumors and acts as a stress-related transcriptional co-activator. Participation of autoimmune responses in the pathophysiology of benign prostatic hyperplasia (PBH) and a corresponding immunosuppressive therapy by TNFalpha antagonists has been recently suggested. Thus, autoAb testing could aid in the diagnosis of BPH patients profiting from such therapy. We generated CRISPR/Cas9 modified HEp-2 LEDGF knock-out (KO) and HEp-2 LEDGF/p75 over-expressing (OE) cells and examined IgG autoantibody reactivity to LEDGF/p75 in patients with prostate cancer (PCa, n = 89), bladder cancer (BCa, n = 116), benign prostatic hyperplasia (BPH, n = 103), and blood donors (BD, n = 60) by indirect immunofluorescence assay (IFA). Surprisingly, we could not detect elevated binding of autoAbs against LEDGF/p75 in cancer patients, but autoAb reactivity to LEDGF/p75 OE cells in about 50% of patients with BPH was unexpectedly significantly increased. Furthermore, a line immunoassay enabling the detection of 18 different autoAbs revealed a significantly increased occurrence of anti-dsDNA autoAbs in 34% of BPH patients in contrast to tumor patients and BD. This finding was confirmed by anti-mitochondrial (mDNA) autoAb detection with the Crithidia luciliae immunofluorescence test, which also showed a significantly higher prevalence (34%) of anti-mDNA autoAbs in BPH. In summary, our study provided further evidence for the occurrence of autoimmune responses in BPH. Furthermore, LEDGF/p75 over-expression renders HEp-2 cells more autoantigenic and an ideal target for autoAb analysis in BPH with a potential therapy consequence.